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Transient activation of human cytomegalovirus lytic gene expression during latency allows cytotoxic T cell killing of latently infected cells.

Krishna BA, Lau B, Jackson SE, Wills MR, Sinclair JH, Poole E - Sci Rep (2016)

Bottom Line: Human cytomegalovirus (HCMV) latency in the myeloid lineage is maintained by repressive histone modifications around the major immediate early promoter (MIEP), which results in inhibition of the lytic viral life cycle.We now show that pharmacological inhibition of histone deacetylases (HDACs) relieves this repression of the MIEP and induces transient expression of the viral lytic immediate early (IE) antigens but, importantly, not full virus reactivation.In turn, these latently infected cells now become targets for IE-specific cytotoxic T cells (CTLs) which are present at high frequency in all normal healthy HCMV positive carriers but would normally be unable to target latent (lytic antigen-negative) cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of Cambridge, Level 5 Laboratories Block, Addenbrooke's Hospital, Hills Road, Cambridge CB2 0QQ.

ABSTRACT
Human cytomegalovirus (HCMV) latency in the myeloid lineage is maintained by repressive histone modifications around the major immediate early promoter (MIEP), which results in inhibition of the lytic viral life cycle. We now show that pharmacological inhibition of histone deacetylases (HDACs) relieves this repression of the MIEP and induces transient expression of the viral lytic immediate early (IE) antigens but, importantly, not full virus reactivation. In turn, these latently infected cells now become targets for IE-specific cytotoxic T cells (CTLs) which are present at high frequency in all normal healthy HCMV positive carriers but would normally be unable to target latent (lytic antigen-negative) cells. This approach of transiently inducing viral lytic gene expression by HDAC inhibition, in otherwise latently infected cells, offers a window of opportunity to target and purge the latent myeloid cell reservoir by making these normally immunologically undetectable cells visible to pre-existing host immune responses to viral lytic antigens.

No MeSH data available.


Related in: MedlinePlus

HDAC4 is increased during latency and targeted by MC1568 to induce lytic gene expression.5 day latent (latency) or control (mock) monocytes were analysed for actin or HDAC4 by Western blot. Samples were run on the same gel and autoradiographs from different exposures were cropped to show the HDAC4 and actin. Blots were analysed using Image J freeware in triplicate. Standard deviations are shown and statistical significance was determined using the Student’s t-test (A). 5 day latent monocytes (latent) or control (mock) monocytes were treated for 24 h with MC1568 or differentiation media. Following 24 h of treatment with MC1568 or 7 day differentiation, cells were stained for IE expression. Data shown (6 replicates) are relative to the level of IE induced following reactivation by differentiation (B) or harvesting for RT-qPCR analysis (C). 5 day latent monocytes were treated with MC1568 and analysed for IE expression after 24 h (Day 1) and then at Day 3, 5 and 7 without removal of MC1568 from the media. Data shown (6 replicates) are relative to the level of IE induced following reactivation by differentiation. Standard deviations are shown and statistical significance was determined using the Student’s t-test (D). 5 day latent monocytes were either untreated (latent) or treated with MC1568 for 24 h (latent plus MC1568) then fixed and stained for IE expression (red) with nuclei shown in blue (E).
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f1: HDAC4 is increased during latency and targeted by MC1568 to induce lytic gene expression.5 day latent (latency) or control (mock) monocytes were analysed for actin or HDAC4 by Western blot. Samples were run on the same gel and autoradiographs from different exposures were cropped to show the HDAC4 and actin. Blots were analysed using Image J freeware in triplicate. Standard deviations are shown and statistical significance was determined using the Student’s t-test (A). 5 day latent monocytes (latent) or control (mock) monocytes were treated for 24 h with MC1568 or differentiation media. Following 24 h of treatment with MC1568 or 7 day differentiation, cells were stained for IE expression. Data shown (6 replicates) are relative to the level of IE induced following reactivation by differentiation (B) or harvesting for RT-qPCR analysis (C). 5 day latent monocytes were treated with MC1568 and analysed for IE expression after 24 h (Day 1) and then at Day 3, 5 and 7 without removal of MC1568 from the media. Data shown (6 replicates) are relative to the level of IE induced following reactivation by differentiation. Standard deviations are shown and statistical significance was determined using the Student’s t-test (D). 5 day latent monocytes were either untreated (latent) or treated with MC1568 for 24 h (latent plus MC1568) then fixed and stained for IE expression (red) with nuclei shown in blue (E).

Mentions: We have previously shown that latent infection of myeloid cells results in major changes in cellular gene expression14 as well as changes in cellular microRNAs15. One cellular microRNA (miRNA) which was identified as being decreased during latency is hsa-miR-20615, which has previously been shown to target HDAC416. Consistent with this, latent infection of monocytes with HCMV did indeed result in elevated levels of HDAC4 (Fig. 1A). Furthermore, the MIEP was found to be heavily associated with HDAC4 during latent infection (Supplementary Figure 1) suggesting a suppressive role for HDAC4 in maintaining latency.


Transient activation of human cytomegalovirus lytic gene expression during latency allows cytotoxic T cell killing of latently infected cells.

Krishna BA, Lau B, Jackson SE, Wills MR, Sinclair JH, Poole E - Sci Rep (2016)

HDAC4 is increased during latency and targeted by MC1568 to induce lytic gene expression.5 day latent (latency) or control (mock) monocytes were analysed for actin or HDAC4 by Western blot. Samples were run on the same gel and autoradiographs from different exposures were cropped to show the HDAC4 and actin. Blots were analysed using Image J freeware in triplicate. Standard deviations are shown and statistical significance was determined using the Student’s t-test (A). 5 day latent monocytes (latent) or control (mock) monocytes were treated for 24 h with MC1568 or differentiation media. Following 24 h of treatment with MC1568 or 7 day differentiation, cells were stained for IE expression. Data shown (6 replicates) are relative to the level of IE induced following reactivation by differentiation (B) or harvesting for RT-qPCR analysis (C). 5 day latent monocytes were treated with MC1568 and analysed for IE expression after 24 h (Day 1) and then at Day 3, 5 and 7 without removal of MC1568 from the media. Data shown (6 replicates) are relative to the level of IE induced following reactivation by differentiation. Standard deviations are shown and statistical significance was determined using the Student’s t-test (D). 5 day latent monocytes were either untreated (latent) or treated with MC1568 for 24 h (latent plus MC1568) then fixed and stained for IE expression (red) with nuclei shown in blue (E).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4835774&req=5

f1: HDAC4 is increased during latency and targeted by MC1568 to induce lytic gene expression.5 day latent (latency) or control (mock) monocytes were analysed for actin or HDAC4 by Western blot. Samples were run on the same gel and autoradiographs from different exposures were cropped to show the HDAC4 and actin. Blots were analysed using Image J freeware in triplicate. Standard deviations are shown and statistical significance was determined using the Student’s t-test (A). 5 day latent monocytes (latent) or control (mock) monocytes were treated for 24 h with MC1568 or differentiation media. Following 24 h of treatment with MC1568 or 7 day differentiation, cells were stained for IE expression. Data shown (6 replicates) are relative to the level of IE induced following reactivation by differentiation (B) or harvesting for RT-qPCR analysis (C). 5 day latent monocytes were treated with MC1568 and analysed for IE expression after 24 h (Day 1) and then at Day 3, 5 and 7 without removal of MC1568 from the media. Data shown (6 replicates) are relative to the level of IE induced following reactivation by differentiation. Standard deviations are shown and statistical significance was determined using the Student’s t-test (D). 5 day latent monocytes were either untreated (latent) or treated with MC1568 for 24 h (latent plus MC1568) then fixed and stained for IE expression (red) with nuclei shown in blue (E).
Mentions: We have previously shown that latent infection of myeloid cells results in major changes in cellular gene expression14 as well as changes in cellular microRNAs15. One cellular microRNA (miRNA) which was identified as being decreased during latency is hsa-miR-20615, which has previously been shown to target HDAC416. Consistent with this, latent infection of monocytes with HCMV did indeed result in elevated levels of HDAC4 (Fig. 1A). Furthermore, the MIEP was found to be heavily associated with HDAC4 during latent infection (Supplementary Figure 1) suggesting a suppressive role for HDAC4 in maintaining latency.

Bottom Line: Human cytomegalovirus (HCMV) latency in the myeloid lineage is maintained by repressive histone modifications around the major immediate early promoter (MIEP), which results in inhibition of the lytic viral life cycle.We now show that pharmacological inhibition of histone deacetylases (HDACs) relieves this repression of the MIEP and induces transient expression of the viral lytic immediate early (IE) antigens but, importantly, not full virus reactivation.In turn, these latently infected cells now become targets for IE-specific cytotoxic T cells (CTLs) which are present at high frequency in all normal healthy HCMV positive carriers but would normally be unable to target latent (lytic antigen-negative) cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of Cambridge, Level 5 Laboratories Block, Addenbrooke's Hospital, Hills Road, Cambridge CB2 0QQ.

ABSTRACT
Human cytomegalovirus (HCMV) latency in the myeloid lineage is maintained by repressive histone modifications around the major immediate early promoter (MIEP), which results in inhibition of the lytic viral life cycle. We now show that pharmacological inhibition of histone deacetylases (HDACs) relieves this repression of the MIEP and induces transient expression of the viral lytic immediate early (IE) antigens but, importantly, not full virus reactivation. In turn, these latently infected cells now become targets for IE-specific cytotoxic T cells (CTLs) which are present at high frequency in all normal healthy HCMV positive carriers but would normally be unable to target latent (lytic antigen-negative) cells. This approach of transiently inducing viral lytic gene expression by HDAC inhibition, in otherwise latently infected cells, offers a window of opportunity to target and purge the latent myeloid cell reservoir by making these normally immunologically undetectable cells visible to pre-existing host immune responses to viral lytic antigens.

No MeSH data available.


Related in: MedlinePlus